The major results of the current study can be summarized as follows. Lafutidine inhibits the vagal afferent signalling of a gastric acid insult, which raises the possibility that lafutidine inhibits acid-induced gastric pain. The ability of lafutidine to decrease intragastric acidity following exposure to excess HCl cannot be explained by its antisecretory activity and is likely to reflect dilution and/or emptying of the acid load into the duodenum.
Vagal afferent signalling of a gastric acid insult was visualized by c-Fos expression in the medullary brainstem, a standard method in functional neuroanatomy to delineate stimulus-evoked activation of neurons [13, 14]. In this way it has previously been shown that exposure of the rat stomach to excess concentrations of HCl stimulates neurons in the brainstem [1, 2]. The appearance of the c-Fos protein was measured 2 h post-HCl, given that the translation of c-fos mRNA into c-Fos protein reaches its maximum between 1 and 3 h post-stimulus . We limited our analysis to the brainstem, because exposure of the rat stomach to HCl failed to induce any c-fos mRNA and c-Fos protein in the dorsal horn of the posterior thoracic spinal cord which receives gastric input via spinal afferent neurons [1, 2]. These data and the ability of chronic bilateral subdiaphragmatic vagotomy to suppress gastric acid-evoked expression of c-fos mRNA  indicate that gastric challenge with HCl is signalled to the brainstem via vagal afferents.
The induction of c-fos mRNA and c-Fos protein in the NTS, the central projection area of vagal afferents, is related to the intragastrically administered HCl concentration [1, 2]. A comparative analysis of the medullary c-Fos induction and gastric damage indicates that the afferent signalling of gastric acid challenge is not directly related to the formation of overt mucosal injury, since c-Fos expression in the NTS can be evoked by HCl concentrations (0.15 – 0.35 M) that do not induce any appreciable macroscopic lesions and cause little histological damage [1, 2]. Because supraphysiological concentrations of HCl (0.15 M or higher) are required to induce c-Fos in the NTS, it has been inferred that only a massive increase in the proton gradient across the acid-tight gastric mucosal barrier is able to drive sufficient protons into the lamina propria where they can excite vagal afferent nerve fibres either directly or indirectly via neuroactive factors released in the tissue . This experimental setup is thus thought to model pathophysiological circumstances where backdiffusion of luminal acid stimulates vagal afferents.
The data of the current study show that lafutidine administered intragastrically at doses (10 and 30 mg/kg) found previously to be gastroprotective [5–6
15reduced the afferent signalling of a gastric acid insult to the NTS. This observation is consistent with the ability of another histamine H2 receptor antagonist, cimetidine, and the proton pump inhibitor omeprazole to reduce gastric acid-evoked expression of c-Fos in the rat brainstem . Since the behavioural reactions to gastric acid challenge indicate that the c-Fos expression in the NTS is a correlate of gastric acid nociception [2, 3] it can be proposed that lafutidine has an inhibitory effect on gastric chemonociception. This antinociceptive effect, however, is unlikely to be explained by the antisecretory activity of lafutidine, given that exposure of the gastric mucosa to excess exogenous acid such as 0.25 M HCl will by itself suppress endogenous acid secretion by a negative feedback mechanism [16–19]. As a consequence, two alternative explanations need to be envisaged.
One explanation that comes to mind is to assume that lafutidine, like cimetidine , interferes with vagal afferent pathways that signal acid challenge of the gastric mucosa to the brainstem. Lafutidine may do so (1) by inhibiting the stimulation of vagal afferent neurons by acid intruding the gastric mucosa, (2) by depressing signal conduction in vagal afferent neurons, (3) by blocking synaptic transmission between the central endings of vagal afferents and NTS neurons and/or (4) by reducing the excitability of the brainstem neurons expressing c-Fos in response to input from the stomach.
Which of these possibilities applies to the effects of lafutidine and cimetidine requires identification of the site of action where H2 receptor antagonists interfere with afferent vagal pathways. It has previously been shown that vagal afferent neurons that signal gastric acid challenge to the NTS are insensitive to the neurotoxic effect of capsaicin  and that the activation of this pathway by acid is independent of prostaglandins . The vagal afferent system inhibited by lafutidine thus appears to be fundamentally different from that of capsaicin-sensitive spinal afferent neurons which are thought to mediate the ability of lafutidine to protect the gastrointestinal mucosa from injury [6–8
15, 21–23]. While vagal afferents appear to be inhibited via a H2 receptor-dependent mechanism, capsaicin-sensitive spinal afferents are activated or sensitized by lafutidine, but not other H2 receptor antagonists, which results in the release of the protective messenger calcitonin gene-related peptide [6–8
15, 21–23]. Prostaglandins may also play a role in the gastroprotective effect of lafutidine against stress .
Another explanation for the inhibitory effect of lafutidine on the gastric acid-evoked c-Fos expression in the brainstem takes into account that lafutidine significantly decreased the acidity of the exogenous gastric acid load. By whatever mechanism lafutidine facilitates the elimination of the gastric acid load, it will also diminish the stimulus for vagal afferent signalling to the NTS. Which of the two mechanisms – direct inhibition of the vagal afferent system or reduction of stimulus strength – is more relevant cannot be decided without electrophysiological characterization of the effect of H2 receptor antagonists on the vagal afferent system.
A direct analysis of the gastroprotective effect of lafutidine in the current study was out of scope because the gastric acid load used here (0.25 M HCl) induced only minimal gastric damage that was indistinguishable from that seen after intragastric administration of saline . As a consequence, gastric acid-induced injury remained unaltered by lafutidine in study 1, although there was a tendency towards suppression of gastric lesion formation. Analysis of the time course of the effect of lafutidine in study 2, however, revealed that lafutidine, unlike vehicle, accelerated the recovery from HCl-induced gastric damage.
The gastroprotective potential of lafutidine is further envisaged from its effect to enhance intragastric pH and to reduce gastric acidity following exposure to an exogenous acid load. Since, as discussed above, endogenous gastric acid secretion is suppressed by an exogenous acid load as high as 0.25 M HCl, the action of lafutidine to decrease gastric acidity may be the result of dilution, neutralization or emptying of the acid load into the duodenum. It has previously been found that exposure of the stomach to supraphysiological concentrations of acid inhibits gastric emptying and causes fluid secretion [2, 9], which in the current study was mirrored by the relatively slow decrease of the gastric volume over the 2 h interval post-HCl.
Lafutidine and cimetidine appeared to accelerate gastric emptying during the period 60 – 120 min post-HCl as indirectly deduced from a decrease in gastric volume that was more pronounced than in vehicle-treated animals. In contrast, the gastric volume immediately after gastric acid loading was significantly increased by lafutidine and cimetidine, which suggests that the secretion of gastric fluid was enhanced. The effect of lafutidine to reduce gastric acidity during the period of 60 – 120 min post-HCl coincided with its apparent effect to accelerate gastric emptying. Apart from this mechanism of action it is also conceivable that lafutidine facilitated gastric acid elimination by dilution with fluid and acid-neutralizing factors. The capacity of lafutidine to stimulate secretory processes other than acid secretion in the upper gut is supported by its effect to increase acid-stimulated duodenal bicarbonate secretion .
Cimetidine shared the effect of lafutidine to inhibit gastric acid-evoked expression of c-Fos in the NTS  and mimicked, in a qualitative manner, the effects of lafutidine to blunt gastric lesion formation, to decrease gastric acidity and to initially enhance and later reduce gastric volume following exposure to an exogenous acid load. The smaller effect of cimetidine, relative to that of cimetidine, is explained by its smaller potency as a H2 receptor antagonist .